tbuckley/cabot-search huggingface dataset:
EXTH-95. UNCOVERING THERAPEUTIC VULNERABILITIES IN MISMATCH REPAIR-DEFICIENT GLIOMAS
Authors: Sangita Pal, Adam Boynton, Ryan Johnston, Naomi Currimjee, Kenin Qian, Mehdi Touat, Charlotte Bellamy, Patricia Ho, David Root, Nicole Persky, Amy Goodale, Jim Berstler, Lisa Miller, Alex Guletsky, Marta Cortes-Cros, Keith Ligon, Rameen Beroukhim, Pratiti Bandopadhayay
Journal: Neuro-Oncology (2022)

Abstract Approximately 25% of recurrent gliomas exhibit hypermutation, which is most often acquired in chemotherapy sensitive gliomas post-treatment with standard of care alkylating agent temozolomide, Intriguingly, nearly all of these recurrent hypermutant tumors harbor deficiencies in mismatch repair (MMR) genes. Unlike other MMR-deficient cancers, hypermutant gliomas do not exhibit detectable microsatellite instability (MSI) at the population level, and do not share similar dependencies. Thus, strategies to therapeutically target MMR-deficient and hypermutant gliomas are urgently needed, and likely to impact many patients. To evaluate vulnerabilities associated with MMR-deficient gliomas, we have generated isogenic MMR-deficient models by systematic ablation of core MMR genes MSH2, MSH6, MLH1, and PMS2, in multiple patient-derived glioma cell lines using a robust all-in-one CRISPR-Cas9 system. We characterized these isogenic MMR-deficient glioma lines in comparison to respective MMR-proficient control line by performing differential gene expression analysis and subsequent gene set enrichment analyses. Our analyses reveal in an unbiased fashion an enrichment of several hallmark gene sets including cell cycle control and DNA repair in the MMR-deficient cell lines. We performed a CRISPR-Cas9 knockout screen in these MMR deficient and hypermutant models and identified candidate genes involved in DNA repair, cell cycle, RNA metabolism and other pathways as preferential dependencies in the MMR-deficient glioma cells. We also screened multiple MMR-deficient isogenic models against the high-throughput drug repurposing (REPO) chemical library and our preliminary results reveal a number of compounds from a variety of drug classes that selectively kill the MMR-deficient glioma cells, indicating that the loss of MMR confers differential dependencies to these small molecule inhibitors. The candidate drugs and genes identified from these high-throughput assays are currently being subjected to further validation, and they possibly will help us identify novel therapeutic strategies or improve existing therapeutic strategies to target recurrent MMR-deficient gliomas.
DDDR-17. CHEMOTHERAPEUTIC STRESS INDUCES NONCANONICAL HIF1Α SIGNALING UNDER NORMOXIA IN GLIOBLASTOMA DRIVING THERAPEUTIC RESISTANCE
Authors: Divya Ventarapragada, Atique Ahmed, Ella Perrault, Cheol Park
Journal: Neuro-Oncology (2022)

Abstract Glioblastoma (GBM) is an aggressive brain tumor with no cure. Standard treatment includes adjuvant, concomitant temozolomide (TMZ) and radiation, but most patients do not survive more than 15 months. Tumor adaptations to temozolomide (TMZ) contribute to glioblastoma (GBM) chemoresistance and recurrence, and hypoxia inducible factor subunit alpha (HIF1α) and downstream effectors are implicated in attenuated therapeutic responses. We have previously shown that TMZ-dependent hijacking of HIF1α signaling under normoxia is independent from physiological hypoxia in the tumor core. Our results further showed that TMZ promotes HIF1α accumulation enhancing stem-like characteristics of GBM cells driving therapy-resistant populations involved in recurrence. However, noncanonical HIF1α signaling mechanisms are not well understood preventing the success of HIF1α inhibitors in GBM thus far. Here, we investigate how TMZ induces HIF1α in an oxygen-independent manner by characterizing the HIF1a binding profile post-therapy. We cultured patient-derived xenograft (PDX) models of GBM in vitro and found HIF1α expression and hypoxia response element (HRE) activity peak 96h post-therapy using western blot and luciferase assays. Our lab also performed a whole-genome CRISPR-Cas9 knockout screen in GBM cells over 28 days to determine genetic drivers of GBM aggression. HIF1α was found to be particularly enriched in this screen, further supporting the role of chronic hypoxia signaling for GBM proliferation (p-value &amp;lt; 0.05). We show TMZ therapy prevents proteasome-dependent degradation through reduced binding of PHD2 and VHL allowing nuclear translocation of HIF1a through western blot and immunocytochemistry. Thus, we performed immunoprecipitation of HIF1α 96h post-therapy for proteomic characterization to establish how TMZ uniquely regulates HIF1α stabilization in the absence of oxygen tension, and these results will be presented. We will subsequently characterize the regulatory role of TMZ-dependent post-translational modifications to HIF1α. Elucidating the molecular mechanisms of therapy-mediated HIF1α regulation can produce better therapeutic targets and increase TMZ efficacy.
PATH-22. CLINICAL FEATURES AND MOLECULAR CHARACTERIZATION OF LEPTOMENINGEAL DISEASE IN PATIENTS WITH HIGH GRADE GLIOMA
Authors: Madison Shoaf, Frances Chow, Joanne Xiu, Michael Glantz, Sonikpreet Aulakh, David Ashley, Eric S Lipp, Giselle Lopez, Ashley Sumrall, Phillip Walker, David Spetzler, Theodore Nicolaides, Katherine B Peters
Journal: Neuro-Oncology (2022)

Abstract INTRODUCTION Leptomeningeal disease (LMD) is a challenging complication of high grade glioma (HGG) and critical questions remain unanswered regarding clinicopathologic risk factors, molecular associations, and optimal treatment. METHODS Patients with molecularly-profiled HGG (Caris Life Sciences; Phoenix, AZ) with LMD at two institutions were included. Medical records were reviewed for clinicopathological characteristics, treatment, and outcome. Kaplan-Meier estimates of patient survival were performed on censored data using Cox’s proportional hazard model. RESULTS 43 patients (male: 33, female: 10; median age: 56 years) were identified, comprising 41 grade 4 (glioblastoma: 38; gliosarcoma: 2; H3K27M diffuse midline glioma: 1) and 2 grade 3 tumors (astrocytoma: 1; pleomorphic xanthoastrocytoma: 1). LMD diagnosed at HGG diagnosis (n=18) versus recurrence (n=22) was associated with longer post-LMD survival [pLMD-OS: 15.3m vs. 4.8m, HR: 0.07, 95% CI: 0.02-0.29, p=0.0004] but similar overall survival [mOS: 15.3m vs. 12.3m; HR: 0.82; 95% CI: 0.36-1.85; p=0.63]. Pathology-diagnosed LMD (n=15) versus MRI-diagnosed LMD (n=26) was associated with longer post-LMD survival [pLMD-OS: 15.4m vs. 5.2m, HR: 14.9, 95% CI: 0.01-0.30, p=0.0004] but similar overall survival [mOS: 17.1m vs. 12.3m; HR: 0.66; 95% CI: 0.3-1.58; p=0.38]. Post-LMD survival was significantly prolonged for supratentorial (n=28) versus infratentorial/spinal (n=4) locations regardless of the diagnostic modality [pLMD-OS: 2.6m vs. 11.3m, HR: 14.4, 95% CI: 2.73-75.7, p=0.0017], and did not significantly differ between symptomatic (n=20) and asymptomatic (n=23) patients [pLMD-OS: 4.8m vs. 11.2m, HR: 1.75, 95% CI: 0.82-3.77, p=0.15). pTERT mutation (81%), EGFR amplification (43%), and MGMT methylation (33%) were prevalent but IDH1 mutation was rare (2.8%). Comparison with a separate glioblastoma cohort (n=1400) suggested more frequent amplification of CHIC2, MDM4, and KDR, higher mutation rates of RUNX1, APC, and RAD51C, colder tumor microenvironment (TME), and lower expression of immune checkpoint-related genes. CONCLUSIONS Clinicopathological characteristics affect post-LMD survival, and cohort comparison suggests molecular and TME differences in LMD-HGG tumors.
DDEL-08. NOVEL COMBINATION THERAPY USING PI3KINASE INHIBITION AND IMMUNE BASED THERAPY FOR THE TREATMENT OF GLIOBLASTOMA MULTIFORME
Authors: Ekokobe Fonkem, Karen newell Rogers, Richard Tobin, Mita Das, Sara Bowen, Ramya Tadipatri, Debbie Healey, Christopher Quarles, John Clark
Journal: Neuro-Oncology (2022)

Abstract Hurdles to effective treatments for glioblastoma multiforme (GBM), the most aggressive form of brain cancer, are resistance to chemotherapy, radiation, and immune checkpoint inhibitors. GBMs, like other “difficult to treat” tumors, often over-utilize the phosphatidyl inositol 3 kinase (PI3K) pathway for survival, cell growth, and cell division, conferring both chemotherapy and radiation resistance. Furthermore, the PI3K pathway has been shown to modulate expression of Major Histocompatibility Complex (MHC) molecules, key components of immune recognition. Based on these observations, we examined the potential of a novel, small molecule PI3K inhibitor GCT.Glio.1 (NPT520-337), which was designed to cross the blood brain barrier, to increase the sensitivity of GBM to immune based therapies and radiation. We hypothesized that targeting PI3K would result in growth arrest and increase cell surface expression of MHC molecules and the PD-L1 checkpoint target. Treatment of GL261 mouse and human U251 GBM cell lines with GCT.Glio.1 caused growth arrest at the G2/M checkpoint, increased the cell surface expression of PD-L1 and MHC class II, and synergized with radiation to cause growth arrest and cell death. Based on these results we designed and performed in vivo animal studies, implanting luciferase-flagged GL261 into the caudate nucleus of C57BL6 mice. We treated the animals orally with GCT.Glio.1 followed 24 hours later by anti-PD1 therapy in repeating cycles. The treatments significantly delayed progression of the tumor when compared to no treatment or either treatment alone, with several animals exhibiting complete regression as measured by MRI and bioluminescence. In conclusion, these results indicate that GCT.Glio.1 may sensitize GBM to immune checkpoint inhibitor and/or radiation therapies. Based on our encouraging pre-clinical data and early results from pre-clinical pharmacology and safety studies, we anticipate filing a pre-Investigational New Drug application (pre-IND) as the next step toward our goal of a clinical trial using this combination therapy.
TMET-38. LOSS OF FUNCTION OF CDK7 IS SYNTHETICALLY LETHAL WITH FATTY ACID OXIDATION INHIBITION IN GLIOBLASTOMA
Authors: Trang Nguyen, Enyuan Shang, Chang Shu, Hasan Akman, Benjamin Izar, Mike-Andrew Westhoff, Georg Karpel-Massler, Jeffrey Bruce, Peter Canoll, Markus Siegelin
Journal: Neuro-Oncology (2022)

Abstract CDK7 has been identified as a potential drug target for glioblastoma (GBM), a highly lethal primary brain tumor. However, resistance to therapy develops quickly, which may be facilitated by drug-induced reprogramming of metabolism. By combination of a transcriptome and metabolite screening analyses followed by carbon tracing (U-13C-Glucose, U-13C-Glutamine and U-13C-Palmitic acid) and extracellular flux analysis we demonstrated that both genetic and pharmacological (YKL-5-124 and THZ1) CDK7 inhibition elicited substantial metabolic reprogramming. Specifically, CDK7i elicited an increase of oxygen consumption rate fueled by enhanced fatty acid oxidation (FAO) manifested by enhanced labeling of citric acid cycle intermediates from palmitic acid. Consistently, the combination treatment of CDK7i inhibitors with blockers of FAO (etomoxir) or cellular respiration (gamitrinib) exerted substantial synergistic growth inhibition in patient derived xenograft as well as neurosphere GBM cultures, which was mainly driven by a collapse of oxidative energy metabolism. In turn, exogenous administration of adenosine triphosphate partially rescued from the cell death induced by the combination treatment. Moreover, the combination treatment activated intrinsic apoptosis through a reduction of both Mcl-1 and Bcl-xL as demonstrated by rescue experiments. Finally, the combined administration of YKL-5-124 and etomoxir extended overall in an orthotopic patient-derived xenograft model of GBM. In summary, these data support that simultaneous targeting of CDK7 and FAO might be a potential novel therapy against GBM.
EPCO-39. ADAPTIVE CONVERGENCE OF METHYLOMES REVEALS EPIGENETIC DRIVERS AND BOOSTERS OF REPEATED RELAPSES IN PATIENT-MATCHED CHILDHOOD EPENDYMOMAS AND IDENTIFIES TARGETS FOR ANTI-RECURRENCE THERAPIES
Authors: Sibo Zhao, Jia Li Li, Huiyuan Zhang, Lin Qi, Yuchen Du, Mari Kogiso, Frank K Braun, Wan-Yee Teo, Holly Lindsay, Patricia A Baxter, Jack M Su, Adekunle Adesina, Sophie Xiao, Clifford Stephan, Stewart G Goldman, Peter J A Davies, Tsz-Kwong Man, Yun Huang, Yongcheng Song, Deqiang Sun, Xiao-Nan Li
Journal: Neuro-Oncology (2022)

Abstract Ependymoma (EPN) is the third most common brain tumor in children and frequently recurs. Here, we report an integrated longitudinal analysis of epigenetic, genetic and tumorigenic changes in 30 patient-matched repeated relapses obtained from 10 pediatric patients to understand the mechanism of recurrences. Genome-wide DNA methylation analysis revealed stable molecular subtypes and convergent epigenetic reprogramming during serial relapses of the 5 RELA and 5 PFA EPNs that paralleled with elevated patient-derived orthotopic xenograft (PDOX) (13/27) formation in the late relapses. Differentially methylated CpGs (DMCs) preexisted in the primary tumors and persisted in the relapses (driver DMCs) were detected, ranging from 51 hypo-methylated in RELA to 148 hyper-methylated DMCs in PFA tumors; while newly acquired DMCs sustained in all the relapses but was absent in the primary tumors (booster DMCs) ranged from 38- 323 hyper-methylated DMCs in RELA and PFA EPNs, respectively. Integrated analysis of these DMC associated DNA methylation regions (DMRs) and RNAseq in both patient and PDOX tumors identified a small fraction of the differentially expressed genes (4.6±4.4% in RELA and 4.5±1.1% in PFA) as regulated by driver DMRs (e.g., up-regulated CACNA1H, SLC12A7, RARA in RELA and HSPB8, GMPR, ITGB4 in PFA) and booster DMRs (including the sole upregulated PLEKHG1 in RELA and NOTCH, EPHA2, SUFU, FOXJ1 in PFA tumors). Most these genes were novel to EPN relapses. Seven DMCs in RELA and 22 in PFA tumors were also identified as potential relapse predictors. Finally, integrating DNA methylation with histone modification identified LSD1 as a relapse driver gene. Combined treatment of a novel inhibitor SYC-836 with radiation significantly prolonged survival times in two PDOX models of recurrent PFA. This high-resolution epigenetic and genetic roadmap of EPN relapse and our 13 new PDOX models should significantly facilitate biological and preclinical studies of pediatric EPN recurrences.
TMET-06. METABOLIC FATE OF OXALOACETATE IN GLIOBLASTOMA
Authors: Omkar Ijare, David Baskin, David Conway, Alan Cash, Kumar Pichumani
Journal: Neuro-Oncology (2022)

Abstract Oxaloacetate (OAA) inhibits the Warburg effect by reducing the glucose-derived lactate levels in human cancer cell lines. In dose ranging IC50 experiments with human hepatocellular carcinoma (HCC) cell lines, OAA inhibited cell growth in the concentration range 1.7 - 2.4 mM. Previously, we showed that in the presence of OAA, [U-13C]glucose derived lactate was reduced by 48.8% in GBM cells. OAA also increased survival in mouse models of GBM. Here, we are showing an evidence for the direct utilization of OAA by Glioblastoma (GBM) cells, using uniformly 13C-labeled oxaloacetate ([U-13C]OAA) as a metabolic tracer. Patient-derived GBM cells were grown in DMEM medium containing 11.0 mM glucose and 2.0 mM glutamine. When cells reached confluency, were treated with 4.0 mM [U-13C]OAA in the above DMEM medium. GC-MS based 13C isotopomer analysis showed that [U-13C]OAA entered the cells and get metabolized to generate distinct 13C-labeled isotopomers of aspartate, citrate, glutamate, malate and fumarate. Upon entering TCA cycle, [U-13C]OAA generated M+4 aspartate (3.0%) and citrate (3.2%), and M+3 glutamate (4.1%). During the first turn of the TCA cycle, it produced M+2 aspartate (13.9%), malate (10.5%), and fumarate (8.8%), and after multiple turns, [U-13C]OAA led to the production of the following 13C-labeled isotopomers: M+2 citrate (21.5%), M+2 glutamate (17.2%), M+1 glutamate (5.6%) and M+4 glutamate (2.2%). In addition, [U-13C]OAA also produced [U-13C]pyruvate (17.0%) through PEPCK/PK/ or malic enzyme which further generated similar amount of [U-13C]lactate (17.0%) and [U-13C]alanine (12%). These data demonstrate that GBM cells utilize [U-13C]OAA to perturb TCA cycle and glycolytic pathways, and support our previous observation that OAA-derived pyruvate lead to an increase in the total pyruvate pool and associated decrease in glycolytic flux including the glucose-derived lactate. OAA may metabolically reduce tumor growth in difficult to treat cancers.
EXTH-07. AN ANTISENSE OLIGONUCLEOTIDE AGAINST TUMOR MICROTUBES IN GLIOBLASTOMA AS A NEW THERAPEUTIC OPTION TO IMPROVE TUMOR SPECIFIC THERAPY
Authors: Sophie Weil, Daniel Domínguez Azorín, Erik Jung, Josie Higgins, Hafeez Mohammed, Jill Reckless, Nigel Ramsden, Peter Keller, David Grainger, Wolfgang Wick, Frank Winkler
Journal: Neuro-Oncology (2022)

Abstract Glioblastomas are known to be highly therapy resistant tumors. The description of tumor microtubes (TMs) as long cellular protrusions that connect glioblastoma cells to a network resistant against all standard therapies advanced the basic biological understanding of the disease and offers a new therapeutic target. An antisense oligonucleotide (ASO) against the production of a protein involved in formation and function of TMs was developed and tested in a xenograft mouse model of human glioblastoma cells. After the implantation of fluorescent tumor cells under a chronic cranial window and the establishment of the tumor, the ASO treatment was started with a continuous micropump application directly into the central nervous system (CNS) and treatment effects were observed in a two-photon microscope. Furthermore, co-treatment with irradiation was tested and tumor size as well as protein expression evaluated in and ex vivo. This led to the conclusion that the setup of combined continuous pump application and two photon microscopy is technically feasible and tolerable. It offers the possibility of applying compounds directly into the CNS under live and longitudinal readouts. The fluorescently tagged ASO was traced in the brain parenchyma over weeks proving a high stability of the compound. In combination with radiotherapy, animals treated with the active ASO against the protein of formation and function of TMs showed a smaller tumor size than animals treated with radiotherapy and the non-targeting ASO. The targeted protein was expressed less in the treated animals then the animals receiving the control ASO and the extend of protein expression correlated with tumor size. Hence, after successful therapies with ASOs in other neurologic diseases and the need for better therapeutic options for glioblastoma patients, the development of an ASO against a protein involved in the TM formation and function offers a new therapeutic option with high translational potential.
STEM-09. NEURAL STEM CELLS OF THE SUBVENTRICULAR ZONE EVOLVE INTO TUMORS AROUND THE RESECTION CAVITY AFTER SURGICAL RESECTION OF GLIOBLASTOMA VIA THE CXCL12/CXCR4 AXIS
Authors: Joo Ho Lee, Xue Li, Hyun Jung Kim, Jeong Ho Lee, Seok-Gu Kang, Hong-Gyun Wu
Journal: Neuro-Oncology (2022)

Abstract Neural stem cells (NSCs) harboring cancer-driving mutations in the subventricular zone (SVZ) have recently been reported as the cells of origin for human GBM. However, the pathological role of residual tumor-initiating NSCs (tNSCs) in the SVZ is undetermined for recurrent tumors after surgical removal of the primary tumor. After the primary resection of GBM, most recurrences still appear as local tumors around the resection cavity (RC). Accordingly, after surgical removal, we hypothesized that the tNSCs in SVZ might be the source for tumor construction at RC as local recurrence. Here, we examined the role of tNSCs in developing local tumors around RC using a mouse model and human tissues. First, cancer-driving mutations, including Trp53, Pten, and Egfr mutations, were introduced to NSCs in SVZ through in vivo electroporation, and the RC was generated in the remote cortex through surgical resection. We found that the NSCs harboring driver mutations migrated specifically to the RC through the aberrant growth of oligodendrocyte-precursor cells (OPCs). Furthermore, the SVZ-originated cells constituted the GBM around the RC at four weeks post-resection in 64% of mouse models. Second, the CXCR4/CXCL12 axis was upregulated through migration from the SVZ and tumor formation at the RC compared with primary tumors without the RC. Blockage of the CXCR4/CXCL12 axis decreased the number of immigrating OPC lineage and switched the differentiation of tNSC to astrocyte lineage. Furthermore, it improved survival rates in vivo. Third, we validated the independent clonal evolution of recurrent tumor from SVZ and the upregulation of CXCR4 axis in the recurrent tumor in a human patient. Taken together, our results reveal that the residual tNCSs harboring cancer-driving mutations in the SVZ can re-construct tumors after surgical resection of GBM. Blocking CXCL12/CXCR4 might be a potential treatment strategy for preventing recurrences after resection.
NIMG-38. 2-HYDROXYGLUTARATE MAGNETIC RESONANCE SPECTROSCOPY IN ADULT BRAINSTEM GLIOMA PATIENTS
Authors: Hirofumi Iwahashi, Hiroaki Nagashima, Kazuhiro Tanaka, Takiko Uno, Mitsuru Hashiguchi, Masahiro Maeyama, Yuichiro Somiya, Masato Komatsu, Takanori Hirose, Tomoo Itoh, Takashi Sasayama
Journal: Neuro-Oncology (2022)

Abstract INTRODUCTION The onco-metabolite, 2-hydroxyglutarate (2HG), is non-invasive biomarker detected by magnetic resonance spectroscopy (MRS), indicating isocitrate dehydrogenase (IDH) mutant glioma. Especially in patients with brainstem lesions where biopsies are at high risk of causing neurological deficit, 2HG-MRS may be useful. Here, we examined the utility of 2HG-MRS for diagnosing IDH mutant adult diffuse brainstem glioma and the ability of prognosis prediction. METHODS Three tesla -MRS (3T-MRS) was performed on patients with radiographically confirmed brainstem tumor. A single-voxel was set in the lesion with reference to the T2 / FLAIR image, and analyzed according to the 2HG-tailored MRS protocol (Philips, Achieva, PRESS, TE 35 ms). All patients underwent biopsy using an intraoperative navigation system (Brain LABTM) or stereotactic biopsy system (Komai’s CT-stereotactic frame) before initial treatment. IDH and H3K27M mutation status were confirmed by immunohistochemistry and DNA sequence. We examined the relationship between 2HG values and patient survival period. RESULTS Ten patients (7 male and 3 female, median age 33.5) underwent 2HG-MRS and biopsy. Four had H3K27M mutation and 4 had IDH mutation (1 R132H, 2 R132S, and 1 R132G). Two had neither H3K27M nor IDH mutation. H3-K27 and IDH mutations were mutually exclusive. All tumor located at pons mainly. There was no significant radiological difference between H3K27M and IDH mutant in conventional MRI sequence. Pearson's chi-square test demonstrated that 2HG concentrations &amp;gt; 1.5 mM were 100% in sensitivity and 83.3% in specificity for IDH mutant glioma (p = 0.0098). The median overall survival was 130.5 months in IDH mutant glioma (n=4) and 25 months in IDH wild-type glioma (n=6), respectively. CONCLUSIONS 3T-MRS was able to detect 2HG in adult brainstem glioma. A noninvasive 2HG-MRS study may be helpful in the diagnosis and predicting prognosis of adult diffuse brainstem glioma patient.